The discovery that enzymes can function in non-aqueous media offers the potential for expanding dramatically the usefulness of enzymes in synthetic chemistry. However, the activity of enzymes in organic media is almost always much lower than that in aqueous solutions. Since many desirable targets for enzyme-catalyzed transformations are insoluble in water, but soluble in organic solvents, low catalytic activity in organic media has thus far limited commercial applications of enzymes in synthetic chemistry. Recent research has shown that lyophilization of serine proteases in a salt matrix activates the enzymes by factors of >1000 for catalysis in organic solvents. The Phase 1 research goals are to refine the methods for preparing salt-activated enzymes, demonstrate the breadth of scope by testing enzymes from different reaction classes, including lipases, metallo-proteases, amidases, dehydrogenases, aminotransferases, and glycosidases, and measure the stability of various salt-activated enzymes in organic solvents. The proposed research would, if successful, result in a broadly applicable method for using enzymes in non-aqueous solvents. As a specific example of commercial importance, salt-activated penicillin amidase will be evaluated in the acylation of two key antibiotic precursors. This reaction will establish the feasibility of an improved method for the enzymatic synthesis of semi-synthetic beta-lactam antibiotics. PROPOSED COMMERCIAL APPLICATIONS: Production of pharmaceutical intermediates and fine chemicals; development of improved biocatalysts